A novel pure component contribution algorithm (PCCA) for extracting components' contribution from severely overlapped signals; an application to UV-spectrophotometric data.

A novel, simple and accurate algorithm capable of extracting the contribution of each component from a mixture signal where the components are completely overlapped was developed. It is based on the development of a coded function which eliminates the signal of interfering components using mean centering as a processing tool; finally the pure contribution of each component is extracted. The algorithm allows the determination of each component as a single one. It was validated by the use of simulated data set of three overlapped signals and tested against simulated random noise. Two fit values were developed and calculated for optimization, one to test that that the absorptivity values of the extracted spectra are within the confidence limits of the slope and the other is the correlation between the pure and extracted spectra. It has been successfully applied to real UV data of binary mixture of Ibuprofen and Paracetamol and ternary mixture of Amiloride hydrochloride, Atenolol and Hydrochlorothiazide in tablets and capsules, respectively. The results were compared to previously reported separation method and no significant difference was found regarding both accuracy and precision.

Spectrofluorometric resolution of closely overlapping drug mixtures using chemometrics methods.

A modified parallel factors analysis (PARAFAC) algorithm with a penalty diagonalization error (PDE), newly proposed by the present authors, was utilized to simultaneously resolve drug mixtures of propranolol (PRO), dipyridamole (DIP) and amiloride (AMI) without any loss of sensitivity. The analyses were performed in aqueous solution. The experimental results demonstrated that the profiles of the spectra and the concentrations could be accurately resolved using the PDE algorithm with a high sensitivity and stable repeatability. That is to say, the closely overlapping problem of the spectra could be easily solved. Furthermore, simultaneous determinations of three kinds of tablets, which contain PRO, AMI and DIP, respectively, were successfully performed with satisfactory results.

Detergent solubilization, functional reconstitution, and partial purification of epithelial amiloride-binding protein.

The amiloride-binding protein from cultured toad kidney cells (A6) was solubilized in 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), functionally reconstituted into liposomes, and partially purified. The specific binding of [3H]methylbromoamiloride ([3H]CH3BrA) was measured in intact A6 epithelia, A6 cell homogenate (H), apical plasma membrane vesicle (V1), and CHAPS-solubilized V1 and on material obtained after affinity chromatography of CHAPS-solubilized plasma membrane vesicles on agarose-immobilized wheat germ agglutinin (WGA). Specific [3H]CH3BrA binding to H, V1, and WGA material reached equilibrium after 10 min. Scatchard analysis of [3H]CH3BrA binding to V1 and WGA material revealed a homogeneous class of binding sites with KD's of 130 and 128 nM, respectively. These KD values were similar to the apparent inhibitory dissociation constant determined from amiloride inhibition of 22Na+ influx in both intact A6 epithelia and V1. The total number of specific binding sites was 4 pmol/mg of V1 protein, which represented a 10-fold enrichment compared to H, and 66.6 pmol/mg of WGA material (a 148-fold enrichment). From association/displacement kinetic studies of specific [3H]CH3BrA binding to V1, the rate constants of association (ka) and dissociation (kd) were calculated to be 3.6 X 10(5) M-1 s-1 and 49.5 X 10(-3) s-1, respectively. These values yield an equilibrium dissociation constant of 138 nM. In solubilized V1 protein, binding activity was enriched approximately 20-fold over H and was markedly dependent upon the relative concentrations of detergent and phospholipid. CHAPS solubilization of V1 resulted in an average 44% recovery of protein with 90% retention of the total number of specific [3H]CH3BrA binding sites. After WGA chromatography 2.7% of the applied protein and 46% of the specific binding sites were recovered.(ABSTRACT TRUNCATED AT 250 WORDS)